bioRxiv Science⌕ Search

Biology subjects

Fernandes, S.

Publications and source records attributed to Fernandes, S..

At least 37 records · Page 2Linked to original sources
bioRxiv2024

Challenges and Efficacy of Astrocyte-to-Neuron Reprogramming in Spinal Cord Injury: In Vitro Insights and In Vivo Outcomes

Traumatic spinal cord injury (SCI) leads to the disruption of neural pathways, causing loss of neural cells, with subsequent reactive gliosis and tissue scarring that limit endogenous repair. One potential therapeutic strategy to address this is to target reactive scar-forming astrocytes with direct cellular reprogramming to convert them into neurons, by overexpression of neurogenic transcription factors. Here we used lentiviral constructs to overexpress Ascl1 or a combination of microRNAs (miRs) miR124, miR9/9*and NeuroD1 transfected into cultured and in vivo astrocytes. In vitro experiments revealed cortically-derived astrocytes display a higher efficiency (70%) of reprogramming to neurons than spinal cord-derived astrocytes. In a rat cervical SCI model, the same strategy induced only limited reprogramming of astrocytes. Delivery of reprogramming factors did not significantly affect patterns of breathing under baseline and hypoxic conditions, but significant differences in average diaphragm amplitude were seen in the reprogrammed groups during eupneic breathing, hypoxic, and hypercapnic challenges. These results show that while cellular reprogramming can be readily achieved in carefully controlled in vitro conditions, achieving a similar degree of successful reprogramming in vivo is challenging and may require additional steps.

neuroscience↗
bioRxiv2024

The homeobox transcription factor Cux1 coordinates postnatal epithelial developmental timing but is dispensable for lung organogenesis and regeneration

Lung epithelial progenitors use a complex network of known and predicted transcriptional regulators to influence early lung development. Here, we evaluate the function of one predicted regulator, Cux1, that we identified from transcriptional regulatory analysis of the SOX9+ distal lung progenitor network. We generated a new Cux1-floxed mouse model and created an epithelial-specific knockout of Cux1 using Shh-Cre (Cux1ShhCre-LOF). Postnatal Cux1ShhCre-LOF animals recapitulate key skin phenotypic features found in prior constitutive Cux1 knockout animals, confirming functionality of the new floxed model. Postnatal Cux1ShhCre-LOF mice displayed subtle alveolar simplification and a transient delay in alveologenesis without persistent lung phenotypes or alterations in lung epithelial cell allocation. Cux1ShhCre-LOF mice developed failure to thrive in their second and third weeks of life due to delayed ileal maturation, which similarly resolves by postnatal day 35. Finally, we challenged Cux1ShhCre-LOF with influenza-mediated lung injury to demonstrate that Cux1ShhCre-LOF mice undergo productive alveolar regeneration that is indistinguishable from WT animals. Together, these findings indicate that epithelial-specific loss of Cux1 leads to transient developmental delays in the skin, lung, and intestine without defects in definitive organogenesis. One-Sentence SummaryDeletion of key DNA binding domains leads to loss of Cux1 function in the lung, intestine, and skin characterized by transient failure to thrive without significant adult disease.

developmental biology↗
bioRxiv2024

An iPSC-derived bio-inspired scaffold modelling the structure and the effects of extracellular matrix in cardiac fibrosis

Cardiac fibrosis occurs following insults to the myocardium and is characterized by the abnormal accumulation of non-compliant extracellular matrix (ECM), which compromises cardiomyocyte contractile activity and eventually leads to heart failure. This phenomenon is driven by the differentiation of cardiac fibroblasts (cFbs) into myofibroblasts and results in changes in ECM biochemical, structural and mechanical properties. The lack of predictive in vitro models of heart fibrosis has so far hampered the search for innovative treatments. Here, we devised a single-step decellularization protocol to obtain and thoroughly characterize the biochemical and micro-mechanical properties of the ECM secreted by activated cFbs differentiated from human induced pluripotent stem cells (iPSCs). We activated iPSC-derived cFbs to the myofibroblast phenotype by tuning basic fibroblast growth factor (bFGF) and transforming growth factor beta 1 (TGF-{beta}1) signalling and confirmed that activated cells acquired key features of myofibroblast phenotype, like SMAD2/3 nuclear shuttling, the formation of aligned alpha-smooth muscle actin (-SMA)-rich stress fibres and increased focal adhesions (FAs) assembly. Next, we used Mass Spectrometry, nanoindentation, scanning electron and confocal microscopy to unveil the characteristic composition and the visco-elastic properties of the abundant, collagen-rich ECM deposited by cardiac myofibroblasts in vitro. Finally, we demonstrated that the fibrotic ECM activates mechanosensitive pathways in iPSC-derived cardiomyocytes, impacting on their shape, sarcomere alignment, phenotype, and calcium handling properties. We thus propose human bio-inspired decellularized matrices as animal-free, isogenic cardiomyocyte culture substrates recapitulating key pathophysiological changes occurring at the cellular level during cardiac fibrosis.

cell biology↗
bioRxiv2023

Moonlight synchronous flights across three western palearctic swifts mirror size dependent prey preferences

Recent studies have suggested the presence of moonlight mediated behaviour in avian aerial insectivores, such as swifts. At the same time swift species also show differences in prey (size) preferences. Here, we use the combined analysis of state-of-the-art activity logger data across three swift species, the Common, Pallid and Alpine swifts, to quantify flight height and activity responses to crepuscular and nocturnal light conditions. Our results show a significant response in flight heights to moonlight illuminance for Common and Pallid swifts, while a moonlight driven response is absent in Alpine swifts. Swift flight responses followed the size dependent altitude gradient of their insect prey. We show a weak relationship between night-time illuminance driven responses and twilight ascending behaviour, suggesting a decoupling of both crepuscular and night-time behaviour. We suggest that swifts optimise their flight behaviour to adapt to favourable night-time light conditions, driven by light responsive and size-dependent vertical insect stratification and weather conditions.

animal behavior and cognition↗
bioRxiv2023

TGF-β induces matrisome pathological alterations and EMT in patient-derived prostate cancer tumoroids

Extracellular matrix (ECM) tumorigenic alterations resulting in high matrix deposition and stiffening are hallmarks of adenocarcinomas and are collectively defined as desmoplasia. Here, we thoroughly analysed primary prostate cancer tissues obtained from numerous patients undergoing radical prostatectomy to highlight reproducible structural changes in the ECM leading to the loss of the glandular architecture. Starting from patient cells, we established prostate cancer tumoroids (PCTs) and demonstrated they require TGF-{beta} signalling pathway activity to preserve phenotypical and structural similarities with the tissue of origin. By modulating TGF-{beta} signalling pathway in PCTs, we unveiled its role in ECM accumulation and remodelling in prostate cancer. We also found that TGF-{beta}-induced ECM remodelling is responsible for the initiation of prostate cell epithelial-to-mesenchymal transition (EMT) and the acquisition of a migratory, invasive phenotype. Our findings highlight the cooperative role of TGF-{beta} signalling and ECM desmoplasia in prompting prostate cell EMT and promoting tumour progression and dissemination

cancer biology↗
bioRxiv2023

YAP signaling regulates the cellular uptake and therapeutic effect of nanoparticles

Interactions between living cells and nanoparticles have been extensively studied to enhance the delivery of therapeutics. Nanoparticles size, shape, stiffness and surface charge have been regarded as the main features able to control the fate of cell-nanoparticle interactions. However, the clinical translation of nanotherapies has so far been limited, and there is a need to better understand the biology of cell-nanoparticle interactions. This study investigated the role of cellular mechanosensitive components in cell-nanoparticle interactions. We demonstrate that the genetic and pharmacologic inhibition of yes-associated protein (YAP), a key component of cancer cell mechanosensing apparatus and Hippo pathway effector, improves nanoparticle internalization in triple-negative breast cancer cells regardless of nanoparticle properties or substrate characteristics. This process occurs through YAP-dependent regulation of endocytic pathways, cell mechanics, and membrane organization. Hence, we propose targeting YAP may sensitize triple negative breast cancer cells to chemotherapy and increase the selectivity of nanotherapy.

bioengineering↗
bioRxiv2023

Importance of genetic architecture in marker selection decisions for genomic prediction

Breeders commonly use genetic markers to predict the performance of untested individuals as a way to improve the efficiency of breeding programs. These genomic prediction models have almost exclusively used single nucleotide polymorphisms (SNPs) as their source of genetic information, even though other types of markers exist, such as structural variants (SVs). Given that SVs are associated with environmental adaptation and not all of them are in linkage disequilibrium to SNPs, SVs have the potential to bring additional information to multi-environment prediction models that are not captured by SNPs alone. Here, we evaluated different marker types (SNPs and/or SVs) on prediction accuracy across a range of genetic architectures for simulated traits across multiple environments. Our results show that SVs can improve prediction accuracy by up to 19%, but it is highly dependent on the genetic architecture of the trait. Differences in prediction accuracy across marker types were more pronounced for traits with high heritability, high number of QTLs, and SVs as causative variants. In these scenarios, using SV markers resulted in better prediction accuracies than SNP markers, especially when predicting untested genotypes across environments, likely due to more predictors being in linkage disequilibrium with causative variants. The simulations revealed little impact of different effect sizes between SNPs and SVs as causative variants on prediction accuracy. This study demonstrates the importance of knowing the genetic architecture of a trait in deciding what markers and marker types to use in large scale genomic prediction modeling in a breeding program. Key messageWe demonstrate potential for improved multi-environment genomic prediction accuracy using structural variant markers. However, the degree of observed improvement is highly dependent on the genetic architecture of the trait.

plant biology↗
bioRxiv2022

Tubulin engineering by semisynthesis reveals that polyglutamylation directs detyrosination

Microtubules, a critical component of the cytoskeleton, carry combinations of post-translational modifications (PTMs), which are critical for the regulation of key cellular processes. Long-lived microtubules, in neurons particularly, exhibit both detyrosination of -tubulin as well as polyglutamylation. Dysregulation of these PTMs results in disease, including developmental defects and neurodegeneration. Despite their importance, the mechanisms governing the emergence of such PTM patterns are not well understood, mostly because tools to dissect the function and regulation of tubulin PTMs have been lacking. Here, we report a chemical method to produce fully functional tubulin carrying precisely defined PTMs within its C-terminal tail. Using a sortase- and intein-mediated tandem transamidation strategy, we ligate synthetic -tubulin tails, which are site-specifically glutamylated to specific extents, to recombinant human tubulin heterodimers. Using microtubules reconstituted with such designer tubulins, we show that polyglutamylation of -tubulin promotes its detyrosination by enhancing the activity of the tubulin tyrosine carboxypeptidase vasohibin/SVBP in a manner dependent on the length of polyglutamyl chains. Moreover, modulating polyglutamylation levels in cells results in corresponding changes in detyrosination. Together, using synthetic chemistry to produce tubulins carrying defined PTMs, we can directly link the detyrosination cycle to polyglutamylation, connecting two key regulatory systems that control tubulin function.

biochemistry↗
bioRxiv2022

Decreased spliceosome fidelity inhibits mTOR signalling and promotes longevity via an intron retention event

Changes in splicing fidelity are associated with loss of homeostasis and ageing1-3, yet only a handful of splicing factors have been shown to be causally required to promote longevity 1-3, and the underlying mechanisms and downstream targets in these paradigms remain elusive. Surprisingly, we found a hypomorphic mutation within RNP-6/PUF60, a spliceosome component promoting weak 3 splice site recognition, which causes aberrant splicing, elevated stress responses, and enhances longevity in Caenorhabditis elegans. Through genetic suppressor screens, we identify a gain-of-function mutation within rbm-39, an RNP-6 interacting splicing factor, which increases nuclear speckle formation, alleviates splicing defects and curtails longevity caused by rnp-6 mutation. By leveraging the splicing changes induced by RNP-6/RBM-39 activities, we uncover a single intron retention event in egl-8/phospholipase C B4 as a key splicing target prolonging life. Genetic and biochemical evidence show that neuronal RNP-6/EGL-8 downregulate mTORC1 signaling to control organismal life span. In mammalian cells, PUF60 downregulation also potently and specifically inhibits mTORC1 signaling. Altogether, our results reveal that splicing fidelity modulates mTOR signaling and suggest a potential therapeutic strategy to delay ageing.

genetics↗
bioRxiv2022

Generation and Maturation of Human iPSC-derived Cardiac Organoids in Long Term Culture

Cardiovascular diseases remain the leading cause of death worldwide; hence there is an increasing focus on developing physiologically relevant in vitro cardiovascular tissue models suitable for studying personalized medicine and pre-clinical tests. Despite recent advances, models that reproduce both tissue complexity and maturation are still limited. We have established a scaffold-free protocol to generate multicellular, beating and self-organized human cardiac organoids (hCO) in vitro from hiPSCs that can be cultured for long term. This is achieved by differentiation of hiPSC in 2D monolayer culture towards cardiovascular lineage, followed by further aggregation on low-attachment culture dishes in 3D. The generated human cardiac organoids (hCOs) containing multiple cell types that physiologically compose the heart, gradually self-organize and beat without external stimuli for more than 50 days. We have shown that 3D hCOs display improved cardiac specification, survival and maturation as compared to standard monolayer cardiac differentiation. We also confirmed the functionality of hCOs by their response to cardioactive drugs in long term culture. Furthermore, we demonstrated that hCOs can be used to study chemotherapy-induced cardiotoxicity. This study could help to develop more physiologically-relevant cardiac tissue models, and represent a powerful platform for future translational research in cardiovascular biology.

cell biology↗
bioRxiv2021

Phenotyping stomatal closure by thermal imaging for GWAS and TWAS of water use efficiency-related genes

Stomata allow CO2 uptake by leaves for photosynthetic assimilation at the cost of water vapor loss to the atmosphere. The opening and closing of stomata in response to fluctuations in light intensity regulate CO2 and water fluxes and are essential to maintenance of water-use efficiency (WUE). However, little is known about the genetic basis for natural variation in stomatal movement, especially in C4 crops. This is partly because the stomatal response to a change in light intensity is difficult to measure at the scale required for association studies. High-throughput thermal imaging was used to bypass the phenotyping bottleneck and assess 10 traits describing stomatal conductance (gs) before, during and after a stepwise decrease in light intensity for a diversity panel of 659 sorghum accessions. Results from thermal imaging significantly correlated with photosynthetic gas-exchange measurements. gs traits varied substantially across the population and were moderately heritable (h2 up to 0.72). An integrated genome-wide and transcriptome-wide association study (GWAS/TWAS) identified candidate genes putatively driving variation in stomatal conductance traits. Of the 239 unique candidate genes identified with greatest confidence, 77 were orthologs of Arabidopsis genes related to functions implicated in WUE, including stomatal opening/closing (24 genes), stomatal/epidermal cell development (35 genes), leaf/vasculature development (12 genes), or chlorophyll metabolism/photosynthesis (8 genes). These findings demonstrate an approach to finding genotype-to-phenotype relationships for a challenging trait as well as candidate genes for further investigation of the genetic basis of WUE in a model C4 grass for bioenergy, food, and forage production. One sentence summaryRapid phenotyping of 659 accessions of Sorghum bicolor revealed heritable stomatal responses to a decrease in light. GWAS/TWAS was used to identify candidate genes influencing traits important to WUE.

plant biology↗
bioRxiv2021

Motor usage imprints microtubule stability in the shaft

Tubulin dimers assemble into dynamic microtubules which are used by molecular motors as tracks for intracellular transport. Organization and dynamics of the microtubule network is commonly thought to be regulated at the polymer ends, where tubulin-dimers can be added or removed. Here we show that molecular motors running on microtubules cause exchange of dimers along the shaft. These sites of dimer exchange act as rescue sites where depolymerising microtubules stop shrinking and start re-growing. Consequently, the average length of microtubules increases depending on how frequently they are used as motor tracks. An increase of motor activity densifies the cellular microtubule network and enhances cell polarity. Running motors leave marks in the shaft serving as traces of microtubule usage to organize the polarity landscape of the cell.

cell biology↗
bioRxiv2021

PI3K Signaling Specifies Proximal-Distal Fate by Driving a Developmental Gene Regulatory Network in Sox9+ Lung Progenitors

The tips of the developing respiratory buds are home to important progenitor cells marked by the expression of SOX9 and ID2. Early in embryonic development (prior to E13.5), SOX9+ progenitors are multipotent, generating both airway and alveolar epithelium, but are selective progenitors of alveolar epithelial cells later in development. Transcription factors, including Sox9, Etv5, Irx, Mycn, and Foxp1/2 interact in complex gene regulatory networks to control proliferation and differentiation of SOX9+ progenitors. Molecular mechanisms by which these transcription factors and other signaling pathways control chromatin state to establish and maintain cell-type identity are not well-defined. Herein, we analyze paired gene expression (RNA-Seq) and chromatin accessibility (ATAC-Seq) data from SOX9+ epithelial progenitor cells (EPCs) during embryonic development. Widespread changes in chromatin accessibility were observed between E11.5 and E16.5, particularly at distal cis-regulatory elements (e.g. enhancers). Gene regulatory network (GRN) inference identified a common SOX9+ progenitor GRN, implicating phosphoinositide 3-kinase (PI3K) signaling in the developmental regulation of SOX9+ progenitor cells. Consistent with this model, conditional ablation of PI3K signaling in the developing lung epithelium in vivo and in lung explants in vitro resulted in an expansion of the SOX9+ EPC population and impaired epithelial cell differentiation. These data demonstrate that PI3K signaling is required for epithelial patterning during lung organogenesis, and emphasize the combinatorial power of paired RNA-Seq and ATAC-Seq in defining regulatory networks in development.

developmental biology↗
bioRxiv2020

Machine learning enabled phenotyping for GWAS and TWAS of WUE traits in 869 field-grown sorghum accessions

Sorghum is a model C4 crop made experimentally tractable by extensive genomic and genetic resources. Biomass sorghum is also studied as a feedstock for biofuel and forage. Mechanistic modelling suggests that reducing stomatal conductance (gs) could improve sorghum intrinsic water use efficiency (iWUE) and biomass production. Phenotyping for discovery of genotype to phenotype associations remain bottlenecks in efforts to understand the mechanistic basis for natural variation in gs and iWUE. This study addressed multiple methodological limitations. Optical tomography and a novel machine learning tool were combined to measure stomatal density (SD). This was combined with rapid measurements of leaf photosynthetic gas exchange and specific leaf area (SLA). These traits were then the subject of genome-wide association study (GWAS) and transcriptome-wide association study (TWAS) across 869 field-grown biomass sorghum accessions. SD was correlated with plant height and biomass production. Plasticity in SD and SLA were interrelated with each other, and productivity, across wet versus dry growing seasons. Moderate-to-high heritability of traits studied across the large mapping population supported identification of associations between DNA sequence variation, or RNA transcript abundance, and trait variation. 394 unique genes underpinning variation in WUE-related traits are described with higher confidence because they were identified in multiple independent tests. This list was enriched in genes whose orthologs in Arabidopsis have functions related to stomatal or leaf development and leaf gas exchange. These advances in methodology and knowledge will aid efforts to improve the WUE of C4 crops.

plant biology↗
bioRxiv2020

Sedimentation rate and organic matter dynamics shape microbiomes across a continental margin

Marine sedimentation rate and bottom-water O2 concentration control the remineralization/sequestration of organic carbon across continental margins; but whether/how they shape microbiome architecture (the ultimate effector of all biogeochemical phenomena), across shelf/slope sediments, is unknown. Here we reveal distinct microbiome structures and functions, amidst comparable pore-fluid chemistries, along ~3 m sediment-horizons underlying the seasonal (shallow coastal) and perennial (deep sea) oxygen minimum zones (OMZs) of the Arabian Sea, situated across the western-Indian margin (water-depths: 31 m and, 530 and 580 m, respectively). Along the perennial- and seasonal-OMZ sediment-cores microbial communities were predominated by Gammaproteobacteria/Alphaproteobacteria and Euryarchaeota/Firmicutes respectively. As a perennial-OMZ signature, a cryptic methane production-consumption cycle was found to operate near the sediment-surface; overall diversity, as well as the relative abundances of simple-fatty-acids-requiring anaerobes (methanogens, anaerobic methane-oxidizers, sulfate-reducers and acetogens), peaked in the topmost sediment-layer and then declined via synchronized fluctuations until the sulfate-methane transition zone was reached. The entire microbiome profile was reverse in the seasonal-OMZ sediment-horizon. We discerned that in the perennial-OMZ sediments organic carbon deposited was higher in concentration, and marine components-rich, so it potentially degraded readily to simple fatty acids; lower sedimentation rate afforded higher O2 exposure time for organic matter degradation despite perennial hypoxia in the bottom-water; thus, the resultant abundance of reduced metabolites sustained multiple inter-competing microbial processes in the upper sediment-layers. Remarkably, the whole geomicrobial scenario was opposite in the sediments of the seasonal/shallow-water OMZ. Our findings create a microbiological baseline for understanding carbon-sulfur cycling across distinct marine depositional settings and water-colum n oxygenation regimes.

microbiology↗
bioRxiv2020

Obesity control by SHIP inhibition requires pan-paralog inhibition and an intact eosinophil compartment

Previously a small molecule SHIP inhibitor, K118, was shown to reverse high-fat diet induced obesity and improve blood glucose regulation in obese mice. K118 treatment was also found to increase the frequency and number of IL-4 producing eosinophils in the visceral fat as well two potent immunoregulatory myeloid cell populations: M2-polarized macrophages and myeloid derived suppressor cells (MDSC) suggesting an immune regulatory mechanism. However, the cell(s) or SHIP paralog that should be targeted to improve metabolic regulation was not defined. Here we extend our understanding of how chemical inhibition of SHIP paralogs improves metabolic regulation during excess caloric intake. Here we compare SHIP inhibitors in an obesity prevention model and find that selective inhibitors for SHIP1 or SHIP2 lack the ability to prevent weight gain and body fat accumulation during increased caloric intake. Surprisingly, only pan-SHIP1/2 inhibitors can prevent diet-induced obesity. We confirm that both SHIP1 and SHIP2 must be targeted by showing that dual treatment with the SHIP1 and SHIP2 selective inhibitors can reduced adiposity caused by excess caloric consumption. We also show that pan-SHIP1/2 inhibitors of two different chemical classes can control diet-induced obesity and improve blood glucose regulation. Intriguingly, we find that both classes of pan-SHIP1/2 inhibitors require an intact eosinophil compartment to prevent diet-related fat accumulation demonstrating pan-SHIP1/2 inhibitors act via the VAT innate immune compartment to control adiposity However, improved blood glucose regulation by pan-SHIP1/2 inhibition is not dependent upon eosinophils, indicating a separate mechanism of action for diet-related loss of glucose regulation.

immunology↗
bioRxiv2019

Metabolically-active obligate aerobes in anoxic (sulfidic) marine sediments

Metabolically-active obligate aerobes are unheard-of in tightly-anoxic environments. Present culture-independent and culture-dependent investigations revealed aerobic microbial communities along two, ~3-meter-long sediment-cores underlying the eastern Arabian Sea oxygen minimum zone, where high H2S disallows O2 influx from the water-column. While genes for aerobic respiration by aa3-/cbb3-type cytochrome-c oxidases and cytochrome-bd ubiquinol oxidase, and aerobic oxidation of methane/ammonia/alcohols/thiosulfate/sulfite/organosulfur-compounds, were present across the cores, so were live aerobic, sulfur-chemolithoautotrophs and chemoorganoheterotrophs. The 8820-years-old, highly-sulfidic, methane-containing sediment-sample from 275 cmbsf of 530 mbsl yielded many such obligately-aerobic bacterial-isolates that died upon anaerobic incubation with alternative electron-acceptors/fermentative-substrates. Several metatranscriptomic reads from this sediment-sample matched aerobic-respiration-/oxidase-reaction-/transcription-/translation-/DNA-replication-/membrane-transport-/cell-division-related genes of the obligately-aerobic isolates, thereby corroborating their active aerobic metabolic-status in situ. Metagenomic and metatranscriptomic detection of perchlorate-/chlorate-reduction genes, plus anaerobic growth of an obligately-aerobic Halothiobacillus isolate in the presence of perchlorate and perchlorate-reducing-consortia, suggested that cryptic O2 produced by perchlorate-respirers could be sustaining obligately-aerobes in this environment.

microbiology↗
bioRxiv2019

Cryptic role of tetrathionate in the sulfur cycle: A study from Arabian Sea oxygen minimum zone sediments

To explore the potential role of tetrathionate in the sulfur cycle of marine sediments, the population ecology of tetrathionate-forming, oxidizing, and respiring microorganisms was revealed at 15-30 cm resolution along two, [~]3-m-long, cores collected from 530- and 580-mbsl water-depths of Arabian Sea, off Indias west coast, within the oxygen minimum zone (OMZ). Metagenome analysis along the two sediment-cores revealed widespread occurrence of the structural genes that govern these metabolisms; high diversity and relative-abundance was also detected for the bacteria known to render these processes. Slurry-incubation of the sediment-samples, pure-culture isolation, and metatranscriptome analysis, corroborated the in situ functionality of all the three metabolic-types. Geochemical analyses revealed thiosulfate (0-11.1 M), pyrite (0.05-1.09 wt %), iron (9232-17234 ppm) and manganese (71-172 ppm) along the two sediment-cores. Pyrites (via abiotic reaction with MnO2) and thiosulfate (via oxidation by chemolithotrophic bacteria prevalent in situ) are apparently the main sources of tetrathionate in this ecosystem. Tetrathionate, in turn, can be either converted to sulfate (via oxidation by the chemolithotrophs present) or reduced back to thiosulfate (via respiration by native bacteria); 0-2.01 mM sulfide present in the sediment-cores may also reduce tetrathionate abiotically to thiosulfate and elemental sulfur. Notably tetrathionate was not detected in situ - high microbiological and geochemical reactivity of this polythionate is apparently instrumental in the cryptic nature of its potential role as a central sulfur cycle intermediate. Biogeochemical roles of this polythionate, albeit revealed here in the context of OMZ sediments, may well extend to the sulfur cycles of other geomicrobiologically-distinct marine sediment horizons.

microbiology↗